Disease resistance in a weaver ant, <Emphasis Type="BoldItalic">Polyrhachis dives</Emphasis>, and the role of antibiotic-producing glands

Parasites represent one of the main threats to all organisms and are likely to be particularly significant for social animals because of the increased potential for intragroup transmission. Social animals must therefore have effective resistance mechanisms against parasites and one of the most important components of disease resistance in ants is thought to be the antibiotic-producing metapleural gland. This gland is ancestral in ants, but has been lost secondarily in a small number of species. It is unknown whether these evolutionary losses are due to a reduction in parasite pressure or the replacement of the gland’s function with other resistance mechanisms. Here we used the generalist entomopathogenic fungus Metarhizium to compare the disease resistance of a species of a weaver ant, Polyrhachis dives, which has lost the metapleural gland, with that of the well-studied leaf-cutting ant Acromyrmex echinatior and two other ant species, Myrmica ruginodis and Formica fusca, all of which have metapleural glands. The P. dives weaver ants had intermediate resistance when kept individually, and similar resistance to A. echinatior leaf-cutting ants when kept in groups, suggesting that the loss of the metapleural gland has not resulted in weaver ants having reduced disease resistance. P. dives weaver ants self-groomed at a significantly higher rate than the other ants examined and apparently use their venom for resistance, as they had reduced resistance when their venom gland was blocked and the venom was shown in vitro to prevent the germination of fungal spores. Unexpectedly, the leaf-cutting ant A. echinatior also had reduced resistance to Metarhizium when its venom gland was blocked. It therefore appears that the evolutionary loss of the metapleural gland does not result in reduced disease resistance in P. dives weaver ants, and that this at least in part may be due to the ants having antimicrobial venom and high self-grooming rates. The results therefore emphasise the importance of multiple, complementary mechanisms in the disease resistance of ant societies.     

Evidence for differential selection and potential adaptive evolution in the worker caste of an inquiline social parasite

Social parasites exploit the socially managed resources of social insect colonies in order to maximise their own fitness. The inquilines are among the most specialised social parasites, because they are dependent on being fully integrated into their host's colony throughout their lives. They are usually relatives of their host and so share ancestral characteristics (Emery's rule). Closely related inquiline-host combinations offer a rare opportunity to study trade-offs in natural selection. This is because ancestral adaptations to a free-living state (e.g. the production of a worker caste) become redundant and may be replaced by novel, parasitic traits as the inquiline becomes more specialised. The dynamics of such processes are, however, unknown as virtually all extant inquiline social parasites have completely lost their worker caste. An exception is Acromyrmex insinuator, an incipient permanent social parasite of the leaf-cutting ant Acromyrmex echinatior. In the present study, we document the size distribution of parasite and host workers and infer how selection has acted on A. insinuator to reduce, but not eliminate, its investment in a worker caste. We show that the antibiotic producing metapleural glands of these parasite workers are significantly smaller than in their host counterparts and we deduce that the metapleural gland size in the host represents the ancestral state. We further show experimentally that social parasite workers are more vulnerable to the general insect pathogenic fungus Metarhizium than are host workers. Our findings suggest that costly disease resistance mechanisms are likely to have been lost early in inquiline evolution, possibly because active selection for maintaining these traits became less when parasite workers had evolved the ability to exploit the collective immune system of their host societies.     

Sanitizing the fortress: protection of ant brood and nest material by worker antibiotics

Social groups are at particular risk for parasite infection, which is heightened in eusocial insects by the low genetic diversity of individuals within a colony. To combat this, adult ants have evolved a suite of defenses to protect each other, including the production of antimicrobial secretions. However, it is the brood in a colony that are most vulnerable to parasites because their individual defenses are limited, and the nest material in which ants live is also likely to be prone to colonization by potential parasites. Here, we investigate in two ant species whether adult workers use their antimicrobial secretions not only to protect each other but also to sanitize the vulnerable brood and nest material. We find that, in both leaf-cutting ants and weaver ants, the survival of the brood was reduced and the sporulation of parasitic fungi from them increased, when the workers nursing them lacked functional antimicrobial-producing glands. This was the case for both larvae that were experimentally treated with a fungal parasite (Metarhizium) and control larvae which developed infections of an opportunistic fungal parasite (Aspergillus). Similarly, fungi were more likely to grow on the nest material of both ant species if the glands of attending workers were blocked. The results show that the defense of brood and sanitization of nest material are important functions of the antimicrobial secretions of adult ants and that ubiquitous, opportunistic fungi may be a more important driver of the evolution of these defenses than rarer, specialist parasites.     

Trail and recruitment pheromones in Camponotus socius (Hymenoptera: Formicidae)

Summary. Worker ants of Camponotus socius employ multicomponent signals during group recruitment to food sources and nest emigrations. The chemical signals consist of trail orientation pheromones that originate from the hindgut. Two components that elicit trail orientation behavior were identified: (2S,4R,5S)-2,4-dimethyl-5-hexanolide and 2,3-dihydro-3,5-dihydroxy-6-methylpyran-4-one. Although both compounds release trail following behavior, in choice tests the former compound is preferred significantly. The major recruitment signal appears to be formic acid discharged from the poison gland. Compounds identified in the Dufour gland secretions do not seem to be involved in the recruitment process.     

Balancing between mutualism and exploitation: the symbiotic interaction between Lasius ants and aphids

Nests of Lasius niger (L.) ants were given varied food regimens to test whether their behaviour towards an aphid partner, Aphis fabae (Scop.), changed with alternative food supplies. Honeydew collection and predation on aphids were measured by video monitoring the movement of ants between their nest and an aphid aggregation. Data collected from the aphid aggregations enabled comparisons between remaining aphid biomass and between the tending intensities of the ants. I tested how ant behaviour was influenced by their access to alternative prey and sugar. The results showed that ants accepted a honey solution as a substitute for the honeydew produced by aphids. Ants not only attended their aphid partners, but also preyed on them. The average predation rate increased eightfold when ants were offered the alternative of sugar, whereas alternative prey had no significant effect. In contrast, ant-tending intensity decreased with alternative sugar whereas alternative prey elicited no effect.     

The origin of the chemical profiles of fungal symbionts and their significance for nestmate recognition in <Emphasis Type="Italic">Acromyrmex</Emphasis> leaf-cutting ants

Cuticular hydrocarbon profiles are essential for nestmate recognition in insect societies, and quantitative variation in these recognition cues is both environmentally and genetically determined. Environmental cues are normally derived from food or nest material, but an exceptional situation may exist in the fungus-growing ants where the symbiotic fungus garden may be an independent source of recognition compounds. To investigate this hypothesis, we quantified the chemical profiles of the fungal symbionts of 18 sympatric colonies of Acromyrmex echinatior and Acromyrmex octospinosus and evaluated the quantitative variation of the 47 compounds in a multivariate analysis. Colony-specific chemical profiles of fungal symbionts were highly distinct and significantly different between the two ant species. We also estimated the relative genetic distances between the fungal symbionts using amplified fragment length polymorphism (AFLP) and correlated these with the overall (Mahalanobis) chemical distances between the colony-specific profiles. Despite the standardized laboratory conditions, the correlations were generally weak, but a statistically significant portion of the total variation in chemical profiles could be explained by genetic differences between the fungal symbionts. However, there was no significant effect of ant species in partial analyses because genetic differences between symbionts tend to coincide with being reared by different ant species. However, compound groups differed significantly with amides, aldehydes, and methyl esters contributing to the correlations, but acetates, alkanes, and formates being unrelated to genetic variation among symbionts. We show experimentally that workers that are previously exposed to and fed with the fungal symbiont of another colony are met with less aggression when they are later introduced into that colony. It appears, therefore, that fungus gardens are an independent and significant source of chemical compounds, potentially contributing a richer and more abundant blend of recognition cues to the colony “gestalt” than the innate chemical profile of the ants alone. Freddie-Jeanne Richard and Michael Poulsen contributed equally to this work.     

Seminal fluid enhances sperm viability in the leafcutter ant Atta colombica

The seminal fluid that accompanies sperm in ejaculates has been shown or suggested to affect sperm competition and paternity success of insects by preventing female remating, inducing oviposition, and forming mating plugs. In Atta leafcutter ants, queens have multiple mates but never remate later in life, although they may live and produce fertilized eggs for several decades. The mating biology and life history of these ants therefore suggests that the major function of seminal fluid is to maximize sperm viability during copulation, sperm transfer, and initial sperm storage. We tested this hypothesis by comparing the viability of testis sperm and ejaculated sperm (mixed with seminal fluid) and found a significant positive effect of seminal fluid on sperm viability. We further quantified this positive effect by adding accessory gland secretion (a major component of seminal fluid) in a dilution series, to show that minute quantities of accessory gland secretion achieve significant increases in sperm viability. Sperm stored by queens for 1 year benefited in a similar way from being exposed to accessory gland compounds after dissection in control saline solution. Our results provide the first empirical evidence that seminal fluid is important for the production of viable ejaculates and that the accessory glands of Atta males—despite their small size—are functional and produce a very potent secretion.     

Predator recognition and evasive behavior by sweat bees, <Emphasis Type="Italic"> Lasioglossum umbripenne </Emphasis>(Hymenoptera: Halictidae), in response to predation by ants, <Emphasis Type="Italic"> Ectatomma ruidum </Emphasis>(Hymenoptera: Formicidae)

Ectatomma ruidum is an abundant soil-nesting Neotropical ant, which displays extensive behavioral flexibility during foraging activities. We studied here one unusual element of their behavioral repertoire: ambush predation. A worker of E. ruidum waits near a nest of a social sweat bee, Lasioglossum umbripenne, lunging at incoming bees, or less frequently, at departing bees. However, bees detected ambushing ants and modified their behavior. Dead ants placed at bees' nest entrances significantly decreased bee activity, indicating that bees recognized dead ants as potential predators. Neither simple black models (square and rectangle) nor olfactory cues had any effect on overall bee activity. A returning bee usually approached her entrance and immediately entered, but if an ant was waiting at the nest, a bee was significantly more likely to abort the first approach flight and then to re-approach the nest on the side opposite the ant's position. As models became increasingly ant-like, returning bees more frequently aborted their first approach flight, expressing other behaviors before entering nests. These behaviors included withdrawal followed by an approach from a different direction; zigzagging flights, either from a distance or close to the entrance or even a close inspection; landing a short distance from the nest, then approaching on foot or waiting for several seconds before entering. Ants responded with effective counter-behaviors. Behavioral flexibility in nest entering/exiting by L. umbripenne and in hunting strategy by E. ruidum shows the complexity of this predator-prey relationship, and illustrates the importance of information processing by both species involved in determining the outcome of the interspecific interaction.     

Chemical trail communication in the amblyoponine species Mystrium rogeri Forel (Hymenoptera, Formicidae, Ponerinae)

Summary. Workers of the amblyoponine species Mystrium rogeri employ trail communication during recruitment to food sources and new nest sites. The trail pheromone originates from a hitherto unknown sternal gland located in the 7th abdominal sternite. The recruiting ant deposits the gland secretions by a special gaster-dragging behavior. The recruitment behavior can be complemented by a rapid vertical body shaking performed by some recruiting ants inside the nest. M. rogeri workers possess a large pygidial gland, the secretion of which elicits a repellent response in other ant species. Received 25 May 1998; accepted 15 June 1998.     

Oily substances from gastral intersegmental glands of the antPachycondyla tridenta (Ponerinae): Lack of pheromone function in tandem running and antibiotic effects but further evidence for lubricative function

Summary Gas chromatographic analyses of volatizable material from gaster intersegmental complex glands ofP. tridentata revealed the presence of linoleic acid, palmitic acid, methyl oleate, and several long-chain hydrocarbons as major constituents, which form an oily fluid mixture. The nest relocation communication ofP. tridentata is tandem running. Tandem following is mediated by pheromones as demonstrated by dummy experiments with isolated gasters and thoraces and with glass balls. The secretions of all gaster complex glands, as well as hind gut contents and metapleural gland secretions, were unable to evoke tandem following. Morphology and position of the glands, lack of pheromonal function, oily properties, low volatility, and lack of antibiotic effects of these secretions strongly suggest a function as lubricants for the ants' gastral segments.     

Defense against parasites by hitchhikers in leaf-cutting ants: a quantitative assessment

Summary A curious behavior in leaf-cutting ants in the genus Atta is the hitchhiking of small minim workers on leaf fragments carried by larger workers. Two functions of these hitchhikers have been proposed: (1) defense of leaf carriers against parasitic flies in the family Phoridae (ant protection hypothesis; Eibl-Eibesfeldt and Eibl-Eibesfeldt 1967) and (2) reduction of transport costs of small minims that collect plant fluids (energy conservation hypothesis; Stradling 1978).We studied hitchhiking behavior in colonies of Atta colombica on Barro Colorado Island, Panama, and found strong evidence in favor of the ant protection hypothesis. Females of Apocephalus attophilus (Diptera: Phoridae) attack leaf carriers of A. colombica and deposit eggs in the head capsules of these ants. Our observations indicate that parasites require leaf fragments to stand on during oviposition, and, as a result, only leaf carriers are susceptible to parasitic attack. The presence of hitchhikers reduces significantly both the time parasites spend on leaf fragments and the probability that they will land in the first place. Results of experimental introductions of parasites and a year of biweekly censuses at ten colonies indicate that leaf-cutting ants adjust the level of hitchhiking to accommodate both daily and seasonal changes in the abundance of parasites.We found little evidence in support of the energy conservation hypothesis. If it is assumed that all minim workers hitchhike back to the nest, our calculations indicate that total transport costs along a foraging trail are reduced by 10% or less. However, our observations indicate that only 50% of returning minim workers hitchhike, and therefore energy savings are actually considerably less than 10%. Leaf-cutting ants in the genera Atta and Acromyrmex are attacked by over 20 species of parasitic phorids. In the discussion we review what is known about these associations and suggest that these parasites have influenced the ecology and evolution of polyethism in leaf-cutting ants. Offprint requests to: D.H. Feener (at his present address)     

Polyethism and the importance of context in the alarm reaction of the grass-cutting ant, Atta capiguara

Leaf-cutting ants exhibit an aggressive alarm response. Yet in most alarm reactions, not all of the ants encountering a disturbance will respond. This variability in behaviour was investigated using field colonies of Atta capiguara, a grass-cutting species. Crushed ant heads were applied near foraging trails to stimulate alarm reactions. We found that minor workers were disproportionately likely to respond. Only 34.7DŽ.8% of ants travelling along foraging trails were minor workers, but 82.1Lj.1% of ants that responded were minors. Workers transporting grass did not respond at all. The alarm response was strongest at the position and time where minors were most abundant. Ants were more likely to respond when they were travelling along trails with low rather than high traffic. Minor workers followed a meandering route along the trail, compared with the direct route taken by foragers. We argue that an important function of minor workers on foraging trails is to patrol the trail area for threats, and that they then play the key role in the alarm reaction.     

Chemical defense of the mint plant, Teucrium marum (Labiatae)

Summary. The mint plant, Teucrium marum (family Labiatae), sometimes called cat thyme, contains two methylcyclopentanoid monoterpenes, dolichodial and teucrein. The former compound is potently anti-insectan. It is repellent to ants (Monomorium pharaonis) and induces preening reflexes in flies (Phormia regina) and cockroaches (Periplaneta americana). Evidence is presented suggesting that dolichodial, which is presumed to be the plant's chief defensive agent, is stored in the tiny epidermal capsules that beset the leaves. It is only when the leaves are injured (and the capsules ruptured) that the leaves become repellent. Teucrein, in contrast, has no anti-insectan potency. It is present predominantly in the leaf buds, unlike dolichodial, which is present mostly in mature leaves. It is argued that teucrein is the storage compound from which dolichodial is generated during leaf development.     

Allometry and the geometry of leaf-cutting in Atta cephalotes

Summary This study considers the relationship of both leg length and the geometry of leaf-cutting to load-size determination by the highly polymorphic leaf-cutting ant Atta cephalotes. A. cephalotes workers anchor on the leaf edge by their hind legs and pivot around them while cutting arcs from leaves. I tested the hypothesis that, for an ant cutting a semicircular leaf fragment, fragment area is determined by a fixed reach while cutting. This reach hypothesis predicts that ants should cut the same fragment-area for at all leaf types. Also, if the radius of the semicircular fragment is proportional to hind leg length, this hypothesis predicts that leaf area should be proportional to hind-leg length squared. The field work was carried out in March–April 1990 and June 1991 in Heredia Province, Costa Rica. I measured hind-leg length for workers of different masses. I then measured leaf-fragment area and mass for workers cutting semicircular fragments from leaves of different densities (mass/area). The logarithmic relationship between ant mass (M _a) and hind-leg length L accelerated negatively (Fig. 1). As a result of this complex allometry, relative leg length (L/M _a ^0.33) increased with ant mass up to a mass of 7.4 mg. Above 7.4 mg, relative leg length decreased. For foragers cutting semicircular fragments, the area cut by an ant of a given size showed no significant difference among leaves of different densities (Fig. 2). Leaf area (A) increased as a function of leg length to the 1.9 power (Fig. 4), an exponent not significantly different from the square function expected if the radius of a fragment is determined by the ant's reach. As a result of this consistent mode of fragment-area determination, the mass of fragments cut by an ant of a given size was significantly greater when cutting denser leaves (Fig. 3) and relative area (A/M _a) cut decreased with increasing ant mass. However, because larger ants generally cut denser leaves (Table 1), the increased density of thick leaves was offset by the reduced relative area cut by the larger ants. Overall, 93% of the foragers cut fragments weighing between 1.5 and 6 times their own body mass (Table 1). Earlier studies found that this broad load-mass range maximized the biomass-transport rate (mass/distance/time) and transport efficiency (mass/distance/energy cost). Thus, A. cephalotes does not solve the problem of matching ant mass and load mass at leaves of different densities with flexibility in the leaf-cutting behavior of individual ants. Instead, individual ants employ a single simple behavioral rule, but workers of different sizes and body proportions tend to cut leaves of different densities.     

A trail pheromone component of the African stink ant, Pachycondyla (Paltothyreus) tarsata Fabricius (Hymenoptera: Formicidae: Ponerinae)

Summary. The African stink ants (Pachycondyla tarsata) lay recruitment trails with secretions from sternal glands. The glandular secretions consist of 10 compounds, 9 of which have been chemically identified. One of the substances, 9-heptadecanone, elicits trail following behavior in P. tarsata workers that have before been stimulated by a sucessful scout ant. Received 7 August 1998; accepted 24 November 1998.     

Quorum sensing, recruitment, and collective decision-making during colony emigration by the ant Leptothorax albipennis

When its nest is damaged, a colony of the ant Leptothorax albipennis skillfully emigrates to the best available new site. We investigated how this ability emerges from the behaviors used by ants to recruit nestmates to potential homes. We found that, in a given emigration, only one-third of the colony's workers ever recruit. At first, they summon fellow recruiters via tandem runs, in which a single follower is physically led all the way to the new site. They later switch to recruiting the passive majority of the colony via transports, in which nestmates are simply carried to the site. After this switch, tandem runs continue sporadically but now run in the opposite direction, leading recruiters back to the old nest. Recruitment accelerates with the start of transport, which proceeds at a rate 3 times greater than that of tandem runs. The recruitment switch is triggered by population increase at the new site, such that ants lead tandem runs when the site is relatively empty, but change to transport once a quorum of nestmates is present. A model shows that the quorum requirement can help a colony choose the best available site, even when few ants have the opportunity to compare sites directly, because recruiters to a given site launch the rapid transport of the bulk of the colony only if enough active ants have been "convinced" of the worth of the site. This exemplifies how insect societies can achieve adaptive colony-level behaviors from the decentralized interactions of relatively poorly informed insects, each combining her own limited direct information with indirect cues about the experience of her nestmates.     

The multifunctional pygidial gland secretion of the Steninae (Coleoptera: staphylinidae): ecological significance and evolution

The pygidial gland secretion of the rove beetle genera Stenus Latreille and Dianous Leach is composed of pyridine and piperidine-derived alkaloids and several terpene compounds. Two-choice bioassays with ants and fish, as well as agar diffusion assays, revealed that the secretion compounds are capable of deterring predators and protecting the beetles from infestation with microorganisms. In addition, the beetles use the secretion for rapid movements on the water surface, a process called skimming. Thus, originally developed to chemically defend the sensitive unprotected abdomen from predator attacks, the secretion of recent Steninae can be designated as multifunctional. Four of the alkaloid compounds occur as different configurational isomers in the secretion. Two-choice tests showed that ants discriminate between stereoisomers of stenusine, while there was no effect visible on bacteria in agar diffusion assays. Furthermore, there are evolutionary trends within the Steninae concerning the secretion composition, as some of the alkaloids primarily occur in phylogenetically basal species, while others are mainly restricted to derived species.     

Waste management in the leaf-cutting ant Acromyrmex echinatior: the role of worker size, age and plasticity

Division of labour is the hallmark of the success of many social animals. It may be especially important with regard to waste management because waste often contains pathogens or hazardous toxins and worker specialisation can reduce the number of group members exposed to it. Here we examine waste management in a fungus-farming, leaf-cutting ant, Acromyrmex echinatior, in which waste management is necessary to protect their vulnerable fungal crop. By marking ants with task-specific paint colours, we found clear division of labour between workers that engage in waste management and those that forage, at least during the fine timescale of the 3-day marking period. This division of labour was influenced by both age and size, with waste management workers tending to be smaller and younger than foragers. The role of preventing contaminated ants from entering the colony was fulfilled mainly by medium-sized workers. When the level of waste was experimentally increased, most of the ants that responded to remove the waste were workers previously engaged in tasks inside the nest rather than external waste workers or foragers. These responding workers tended to be young and medium-sized. Surprisingly, the responding ants were subsequently able to revert back to working within the fungus garden, but the probability of them doing so depended on their age and the length of time they were exposed to waste. The results demonstrate the importance of division of labour with regard to waste management in A. echinatior and show that this is adaptable to changing needs.     

Three beetles—three concepts. Different defensive strategies of congeneric myrmecophilous beetles

Myrmecophiles, i.e., organisms associated with ants live in a variety of ecological niches in the vicinity or inside ant colonies and employ different strategies to survive ant encounters. Because different niches are characterized by different encounter rates with host ants, strategies used to avoid ant aggressions should depend on these niches. This hypothesis was studied with three rove beetle species of the genus Pella, which are myrmecophiles of the ant Lasius fuliginosus and the non-myrmecophilous relative Drusilla canaliculata. Behavioral tests in the field revealed that Pella species are better adapted to interactions with ants than D. canaliculata, but that they use species-specific strategies in ant interactions. Pella cognata and Pella funesta avoid encounters with ants by swift movements. Chemical analyses of the defensive tergal gland secretions showed that P. cognata has replaced the aggression inducing undecane by the behaviorally neutral tridecane. P. funesta repels the ants by releasing the panic alarm pheromone sulcatone from its tergal gland resulting in an “ant free space” around the beetles. Finally, Pella laticollis uses a specific and unique appeasing behavior. Behavioral and chemical data did not reveal any indication for the mimicry of the ants' cuticular hydrocarbon profiles by any of the beetle species. It is discussed that the employed strategies correlate with the ecological niches of the beetles. P. cognata and P. funesta are living along ant trails with ample space to escape and the employed strategies are probably sufficient to escape from dangerous conflicts. In contrast, P. laticollis lives in refuse areas of ant nests with frequent ant encounters, and its appeasement strategy allows it to stay at the encounter site.     

Experimental conversion of colony social organization by manipulation of worker genotype composition in fire ants (<Emphasis Type="Italic">Solenopsis invicta</Emphasis>)

Previous studies have shown that colony social organization in Solenopsis invicta is under strong genetic control. Colonies containing some proportion of workers with the Bb or bb genotypes at the gene Gp-9 display polygyne social organization (multiple reproductive queens per colony), whereas colonies with only BB workers express monogyne organization (single reproductive queen per colony). The hypothesis that the presence of workers bearing the b allele confers the polygyne social phenotype on a colony leads to the prediction that social organization can be manipulated by experimentally altering frequencies of adult workers bearing this allele. We did this by replacing queens in colonies of each social form with single queens of the alternate form, which differ in Gp-9 genotype. As worker Gp-9 genotype compositions changed, experimental colonies switched to the alternate social organization. These switches occurred when frequencies of workers with the b allele passed an identifiable threshold, such that colonies with fewer than 5% such workers behaved like monogyne colonies and those with more than 10% behaved like polygyne colonies. Our data thus confirm the prediction that colony social organization in this ant can be altered by manipulating adult worker genotype compositions, and thereby support the hypothesis that the expression of polygyny requires the presence of adult workers bearing the b allele at Gp-9.